| Objective:To investigate the effects of high glucose on proliferation and oxidative stress of human dental pulp cells(HDPCs),in order to explore the mechanism of high glucose conditions on dental pulp.Methods:1、Isolation,culture and identification of human dental pulp cells in vitro.2、The experimental group:The HDPCs were divided into four groups:the glucose concentration of 5.5 mmol/L was set as low glucose group,25mmol/L as normal group,50 mmol/L as high glucose group,osmotic pressure was equal to that of the 50 mmol/L group and glucose concentration was equal to that of the 5.5 mmol/L group as hypertonic group.3、Different concentrations of glucose culture HDPCs,after 1、3、5days,CCK-8 kits were used to detect the proliferation condition,draw the growth curve.4、Cells of each experimental group were treated with2,7-dichlorofuorescin diacetate(DCFH-DA),a kind of Reactive oxygen species(ROS)detection agent,fluorescence intensity in HDPCs was observed by fluorescence microscope,and intracellular ROS level was detected by multiscan spectrum.The content of malondiadedehyde(MDA)in cells treated with different concentrations of glucose was detected by enzyme labeling method,and the activity of superoxide dismutase(SOD)was measured by Water-soluble tetrazolium method(WST-1).5、Cells of each experimental group were incubated with the fluorescent probe JC-1,and the changes in Mitochondrial transmembrane potential(MMP)of HDPCs were expressed by flow cytometry analysis.Results:1、The pulp cells cultured by modified tissue block method showed long spindle shape and good growth.HDPCs anti-keratin staining was negative,no staining was found in the cytoplasm,anti-vimentin was positive,and brown-yellow positive particles were visible in the cytoplasm.2、Compared with the normal group,high concentration of glucose culture dental pulp cells,after 1,3 and 5 days,the proliferation of HDPCs was slow and the cell activity was low,showing an obvious inhibitory effect.3、The levels of reactive oxygen species(ROS)and malondialdehyde(MDA)increased significantly after being exposed to glucose concentration of 50 mM for 72 h,while the activity of superoxide dismutase(SOD)decreased.4、The detection of mitochondrial membrane potential showed that the fluorescence value of the high-glucose group was higher than that of the other experimental groups,with a statistical difference(P<0.05).Conclusion:1、The high glucose can reduce the activity of dental pulp cells,cell proliferation has an obvious inhibitory effect.2、The high glucose environment can cause the decrease of mitochondrial membrane potential of HDPCs,significantly increase ofROS and MDA levels,and decrease of SOD content,which can induce the generation of oxidative stress in pulp cells. |
